LOCUS       sequence-430815-        6554 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The DuET construct encodes the named human GPCR engineered with a 
            cleaved hemagglutinin signal peptide, a 5' FLAG and a 3' 1D4 epitope
            tag optimized for expression in mammalian cells and proteomics 
            studies..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6554)
  AUTHORS   Dahl L, Kotliar IB, Bendes A, Dodig-Crnkovic T, Fromm S, Elofsson A,
            Uhlen M, Sakmar TP, Schwenk JM
  TITLE     Multiplexed selectivity screening of anti-GPCR antibodies.
  JOURNAL   Sci Adv. 2023 May 3;9(18):eadf9297. doi: 10.1126/sciadv.adf9297. 
            Epub 2023 May 3.
  PUBMED    37134173
REFERENCE   2  (bases 1 to 6554)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2023-05-3"; volume: "9(18):eadf9297. doi"; pages: " 
            10.1126/sciadv.adf9297. Epub 2023 May 3"
COMMENT     Sequence Label: GPR160-DuET
FEATURES             Location/Qualifiers
     source          1..6554
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     485..504
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(723..827)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(902..921)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1093..1472
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1473..1676
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1627..1647
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1721..1740
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1721..1739
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3003..3020)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3009..3233
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3279..3707
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3366..3385)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3576..3597
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3716..3736)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3721..4050
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3901..4036
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3963..3982
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4117..4911
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4171..4190)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4781..4800
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5085..5206
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5122..5141)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5176..5195
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5255..5271)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5255..5271)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5268..5290)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5279..5295
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5303..5309,5310..5327,5328..5333))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5348..5369
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5486..5503)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5657..6245)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5737..5756)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6416..6554,1..653,654..722))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
       61 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
      121 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
      181 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
      241 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
      301 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
      361 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
      421 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
      481 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
      541 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
      601 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
      661 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
      721 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
      781 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
      841 aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg tgcactctca
      901 gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct tgtgtgttgg
      961 aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca
     1021 tttgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca
     1081 gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
     1141 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
     1201 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
     1261 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
     1321 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
     1381 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
     1441 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
     1501 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
     1561 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
     1621 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct
     1681 ggctaactag agaacccact gcttactggc ttatcgaaat taatacgact cactataggg
     1741 agacccaagc tggctagcgt ttaaacttaa gcttagcgcc accatgaaga cgatcatcgc
     1801 cctgagctac atcttctgcc tggtattcgc cgactacaag gacgatgatg acgccagcat
     1861 cgatatgact gccctgagca gtgagaattg ttcattccag taccagctta gacagacaaa
     1921 tcaaccactc gacgtgaact acctgctttt cctcataatt ctgggaaaga ttcttttgaa
     1981 catccttaca ctgggtatgc gccggaaaaa cacatgccag aactttatgg agtacttctg
     2041 cattagcttg gcctttgtcg acttgctcct gttggttaat atctctatca tcctgtactt
     2101 ccgggacttc gttctcctta gtatcaggtt cactaagtat cacatttgtt tgttcaccca
     2161 aatcatatcc tttacatatg gattcctcca ctacccagtt tttctcaccg cttgtatcga
     2221 ctattgtctg aactttagca aaacgaccaa actctcattc aagtgccaga aactctttta
     2281 tttctttacc gttatactga tttggatatc cgtactggcc tatgtgctgg gtgatcccgc
     2341 catataccag tccctgaagg cccagaatgc ctattccagg cactgcccct tttacgtgag
     2401 tattcagagt tactggctct cctttttcat ggtgatgatt ctgtttgttg ctttcatcac
     2461 gtgctgggag gaagtgacca cactggtcca agctattcgg atcacctctt acatgaatga
     2521 gaccattctt tatttcccat ttagctccca cagcagctat actgtgcgga gcaaaaaaat
     2581 cttccttagt aagctgatcg tgtgtttcct gagcacctgg ctgcccttcg tgcttctcca
     2641 ggtgataatt gtgctgctga aggtgcagat ccccgcgtac attgagatga atattccatg
     2701 gctgtacttt gtgaactctt ttttgatcgc tactgtctat tggttcaatt gccacaagct
     2761 caaccttaag gatattggtc tgccccttga ccccttcgtg aattggaaat gctgttttat
     2821 tcctcttaca atcccgaatc tcgaacagat cgagaagccg atatccatca tgatctgtat
     2881 cgataccggt ggacgcaccg gtgaattcga tgaggccagc accaccgtgt ctaagaccga
     2941 gacttctcag gtggcccctg cctaactcga gtctagaggg cccgtttaaa cccgctgatc
     3001 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     3061 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     3121 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     3181 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     3241 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     3301 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3361 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3421 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3481 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3541 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3601 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3661 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     3721 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3781 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3841 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3901 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3961 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     4021 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     4081 ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac
     4141 gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca
     4201 atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt
     4261 gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg
     4321 tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga
     4381 agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct
     4441 cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg
     4501 gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg
     4561 gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc
     4621 gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat
     4681 ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac
     4741 tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt
     4801 gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct
     4861 cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc
     4921 tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca
     4981 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     5041 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     5101 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     5161 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     5221 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     5281 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     5341 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     5401 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     5461 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     5521 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     5581 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5641 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5701 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5761 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     5821 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     5881 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     5941 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     6001 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     6061 tcttgaagtg gtggcctaac tacggctaca ctagaagaac ggtatttggt atctgcgctc
     6121 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6181 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6241 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6301 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6361 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6421 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6481 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     6541 ctgcaatgat accg
//
